C. trachomatis is the most common sexually transmitted bacterial pathogen in the US. Chlamydia infections appear to have adverse effects on women's health including complications of pregnancy, pelvic inflammatory disease, infertility, and cervical infections. The objectives of this project are to define the epidemiology, risk factors, transmission kinetics and pathogenesis of Chlamydia trachomatis infections in different population settings and different disease states using new molecular amplification assays. To address this objective, we implemented non-invasive screening with molecular assays for C. trachomatis and documented high rates of infection in sexually active adolescents in diverse cultural settings in six countries. Prevalence rates of chlamydia ranged from 3.8% in rural villages in Zimbabwe and Uganda, 12.5% in St. Petersburg, Russia, 11.5% in Lima, Peru, 16.7% in Fuzhou, China, and over 20% in Baltimore, USA. The documented high rates of chlamydia in these countries raises serious concerns about the resurgence of STDs that may reflect a rise in high-risk behavior which may subsequently lead to further HIV transmission. In a study of adolescent males in the US, we determined the cost effectiveness of chlamydia screening on admission to detention centers. Chlamydia prevalence in the population was 4.8% and the average number of female sexual partners per infected male was 1.6. We determined that universal screening by nucleic acid amplified test was the most cost- effective strategy preventing 37 more cases of pelvic inflammatory disease and three more cases of epididymitis than selective screening. With the implementation of universal screening we have performed several studies comparing self-administered vaginal swabs, cervical swabs, and first-catch urines for the diagnosis of chlamydial infections in women. Specimens from 22,517 15-to-25-year-old asymptomatic women attending clinics in nine different centers in the U.S. were evaluated. Overall, chlamydia prevalence was 13%. Results with a self-administered vs. a clinician-collected vaginal swab were equivalent and were at least as good as results with first-catch urines and cervical swabs. Nucleic acid amplified testing sensitivity with vaginal swabs was slightly higher (93%) than with cervical swabs (91%) or first-catch urines (81%) or culture of cervical swabs (84%). These studies demonstrate that vaginal swabs are appropriate specimens for diagnosing chlamydial genital tract infection by amplified diagnostic assays. Non-invasive screening for chlamydia can be applied in different clinical and non-clinical sites thereby increasing the size of the population to be screened Utilizing the same principles for DNA detection, we have developed a molecular diagnostic assay for the detection of C. pneumoniae, a frequent causes of community-acquired pneumonia and which contributes significantly to the morbidity of individuals suffering from respiratory disease. We developed a real-time PCR for C. pneumoniae demonstrating an analytical sensitivity of detection between 4 and 0.4 infection-forming units per PCR reaction. A total of 355 samples were tested for validation of the assay and results were compared to nested PCR. Overall, the real-time PCR for C. pneumoniae had a sensitivity of 89% and a specificity of 99.3%, demonstrating its utility as an accurate, high-throughput detection method for detection of C. pneumoniae infection. Future investigations will utilize this highly sensitive, rapid and specific assay for detection of C. pneumoniae in respiratory and atherosclerotic lesions.